Scientific Report 2003-2004 - Cleveland Clinic Lerner Research ...

ORTHOPAEDICBIOLOGY ANDBIOENGINEERINGTHE HASCALLLABORATORYPROJECT SCIENTISTSAnthony Calabro, Ph.D.Csaba Fulop, Ph.D.Aimin Wang, Ph.D.POSTDOCTORAL FELLOWSMark Lauer, Ph.D.Durba Mukhopadhyay, Ph.D.COLLABORATORSAnthony Day, D. Phil. 1Carol de la Motte, Ph.D. 2Edward Maytin, M.D. 3Judy Mack, Ph.D. 3Antonietta Salustri, Ph.D. 4Scott Strong, M.D. 2Markku Tammi, M.D. 5Raija Tammi, M.D. 51MRC Immunochemistry Unit,Univ. of Oxford, Oxford, UK.2Dept. of Colorectal Surg. andImmunol. Res., CCF3Dept. of BiomedicalEngineering, CCF4Dept. of Cell Biol., 2 nd Univ. ofRome, Rome, Italy5Dept. of Anatomy, Univ. ofKuopio, Kuopio, FinlandOur laboratory focuses on the structure,function and metabolism of proteoglycansand hyaluronan in connective tissues.Proteoglycans are specialized proteins containingone or more covalently bound glycosaminoglycanchains: chondroitin/dermatan sulfate, keratan sulfate,heparin/heparan sulfate, or a combination of types.Glycosaminoglycan chains comprisepolymers of repeating disaccharide motifs that(except for hyaluronan) contain sulfoestersubstituents along their carbohydrate backbones.These highly negatively charged glycosaminoglycansendow the parent proteoglycans with a widerange of structures and functions. A briefdescription of proteoglycans currently under studyfollows.The major proteoglycan of cartilages isaggrecan; its core protein is ~200,000 Da molecularweight, with >150 chondroitin sulfate and keratansulfate chains attached. The mature proteoglycan(>2 million Da molecular weight) is the majorcomponent of cartilage extracellular matrix andenables this tissue to resist compressive load withminimal deformation.Biosynthesis and catabolism of aggrecan areclosely regulated, interdependent processesessential for normal tissue function, both duringlong-bone development on cartilage templates andin maintenance of articular cartilages that absorbshock on the ends of mature long bones. Abnormalbiomechanical load on cartilages, e.g., afterinjury to the knee’s cruciate ligaments, can adverselyalter aggrecan metabolism in the tissue and causematrix degeneration that eventually leads to clinicalosteoarthritis. Our goal is to use model chondrocyteor cartilage tissues to study processes thatregulate the amount and fine structure of aggrecansynthesized in normal and pathological situationsand to determine how aggrecan synthesis andcatabolism are regulated.The macromolecule hyaluronan is notsynthesized on a core protein, but rather at or nearthe cell surface, with the growing polymer beingThe Department of Biomedical EngineeringHyaluronan and Proteoglycan BiochemistryForm the Basis of Tissue EngineeringResearch Programextruded into the extracellular space. Hyaluronanconsists of repeating disaccharides of glucuronicacid and N-acetylglucosamine and reaches molecularweights >10 million Da.Hyaluronan is nearly ubiquitous throughoutthe vertebrates and is also synthesized by somebacteria. It serves as a scaffold for the extracellularmatrix in many connective tissues and has a vitalrole in tissue morphogenesis. Despite its prevalenceand deceptively simple structure, details of itssynthesis and metabolic regulation remainenigmatic.We have studied hyaluronan’s role in severalsystems. In cartilage, hyaluronan forms a filamentouslattice structure in the matrix to which aggrecanmolecules are anchored. Its metabolism is closelycorrelated with that of aggrecan and involves cellsurfacehyaluronan receptors as part of catabolism.In the final stages preceding ovulation, the oocytedirects the adjacent cumulus cell population tosynthesize and organize a similar hyaluronan lattice,surrounding itself with an extracellular matrix thatplays a vital role in subsequent fertilization. Inkeratinizing epithelia (e.g., skin or oral mucosa), thecells synthesize hyaluronan, which forms a matrixaround the basal and the differentiating suprabasalcells. These epithelia continuously regenerate andundergo terminal differentiation in the keratinizinglayers; thus hyaluronan must also be catabolizedcompletely within the tissue. Hyaluronan producedby colon or lung smooth muscle cells in responseto viral stimuli is organized in macrostructures thatengage and activate mononuclear leukocytes. Thismechanism is central in the pathogenesis ofinflammatory bowel disease and asthma. Thesecells, as well as mesangeal cells, synthesize similarstructures in response to elevated glucose concentrationsas in uncontrolled diabetes, suggesting thatabnormal hyaluronan matrices synthesized inresponse to elevated glucose are also involved invascular and diabetic pathologies. We are studyingthe mechanisms of both biosynthesis and catabolismof hyaluronan in these systems.Calabro, A., Oken, M.M., Hascall, V.C., and A.M. Masellis (2002) Characterization of hyaluronan synthaseexpression and hyaluronan synthesis in bone marrow mesenchymal progenitor cells: predominantexpression of HAS1 mRNA and up-regulated hyaluronan synthesis in bone marrow cells derived frommultiple myeloma patients. Blood 100:2578-2585.Vincent C. Hascall, Ph.D.See the website“Science ofHyaluronan”http://www.glycoforum.gr.jpKnepper, M.A., Saidel, G.M., Hascall, V.C., and T. Dwyer (2003) Concentration of solutes in the renal innermedulla: interstitial hyaluronan as a mechano-osmotic transducer. Am. J. Physiol. Renal Physiol.284:F433-F446.Fulop, C., Szanto, S., Mukhapadhyay, D., Bardos, T., Kamath, R., Day, A., Salustri, A., Hascall, V.C.,Glant, T., and K. Mikecz (2003) Impaired cumulus mucification and female sterility in tumor necrosisfactor-induced protein-6-deficient mice. Development. 130:2253-2261.Mack, J.A., Abramson, S.R., Ben, Y., Coffin, J.C., Rothrock, J.K., Maytin, E.V., Hascall, V.C., Largman,C., and E.J. (2003) Hoxb13 knockout adult skin exhibits high levels of hyaluronan and enhancedwound healing. FASEB J. 2003 May 20 [Epub ahead of print].de la Motte, C.A., Hascall, V.C., Drazba, J., Bandyopadhyaya, S.K., and S. Strong (2003) Mononuclearleukocytes bind to specific hyaluronan structures on colon mucosal smooth muscle cells treated withpolyinosinic acid:polycytidylic acid. Inter-alpha-trypsin inhibitor is crucial to structure and function. Am.J. Pathol. 163:000-000 (in press).36